Abstract

A loss-of-function mutation in the gene parkin causes a common neurodegenerative disease that may be caused by mitochondrial dysfunction. Glutathione S-transferase Omega (GSTO) is involved in cell defense mechanisms, but little is known about the role of GSTO in the progression of Parkinson disease. Here, we report that restoration of Drosophila GSTO1 (DmGSTO1), which is down-regulated in parkin mutants, alleviates some of the parkin pathogenic phenotypes and that the loss of DmGSTO1 function enhances parkin mutant phenotypes. We further identified the ATP synthase β subunit as a novel in vivo target of DmGSTO1. We found that glutathionylation of the ATP synthase β subunit is rescued by DmGSTO1 and that the expression of DmGSTO1 partially restores the activity and assembly of the mitochondrial F(1)F(0)-ATP synthase in parkin mutants. Our results suggest a novel mechanism for the protective role of DmGSTO1 in parkin mutants, through the regulation of ATP synthase activity, and provide insight into potential therapies for Parkinson disease neurodegeneration.

Up-regulation of DmGSTO1 in muscle decreases tubulin accumulation and UPR activation in park1 mutants. Shown are representative images of flight muscle stained with anti-α-tubulin antibody and phalloidin. A, α-tubulin accumulates in park1 mutant muscle. The accumulation of tubulin in the park1 mutant was suppressed by the muscle-specific up-regulation of DmGSTO1A. B, the levels of actin filaments in muscle did not change in any mutants as visualized with phalloidin. C, the increased levels of α/β-tubulin in park1 mutants were rescued by DmGSTO1A expression. Error bars, S.D. The experimental significance was determined by one-way ANOVA (**, p < 0.05; *, p < 0.01). D, tubulin levels were higher in the double mutants than in the park1 single mutants. Error bars, S.D. The significance was determined by one-way ANOVA (***, p < 0.01; **, p < 0.05). E, mRNA levels of male thoraces were determined by real-time PCR analysis. Relative amounts of tubulin mRNA were unchanged in all mutants. F, Western blot analysis of phosphorylated and total eIF2α. G, increased HSPs in park1 mutants were decreased with the up-regulation of DmGSTO1A. F and G, error bars, S.D. The significance was determined by one-way ANOVA (*, p < 0.05). β-Actin was used as a loading control. Experiments were performed in triplicate.

The catalytic activity of DmGSTO1 is critical for rescue of the defective phenotypes of the park1 mutant.A, expression of DmGSTO1AC31A in the park1 mutant background did not suppress the collapsed thorax (n > 200) and downturned wing (n > 160) phenotypes of parkin mutants. Error bars, S.D. The experimental significance was determined by one-way ANOVA (*, p < 0.05). B, Western blot analysis of adult thorax extracts using anti-β-tubulin antibodies. DmGSTO1AC31A expressed in the park1 mutants did not rescue the tubulin accumulation phenotypes. Error bars, S.D. The significance was determined by one-way ANOVA (**, p < 0.05; *, p < 0.01). β-Actin was used as a loading control. Experiments were performed in triplicate.